From Left: Lee Jialing, Dr Melissa Lin, Dr Steve Oh
Youshan Melissa Lin 1*, Jessica Fang Yan Lim 1*, Jialing Lee 1, Mahesh Choolani 2, Jerry Kok Yen Chan 2,3, Shaul Reuveny 1, Steve Kah Weng Oh 1
1 Bioprocessing Technology Institute, Agency for Science, Technology and Research (A*STAR), Singapore
2 Experimental Fetal Medicine Group, Department of Obstetrics and Gynaecology, Yong Loo Lin School of Medicine, National University Health System, Singapore
3 Department of Reproductive Medicine, KK Women’s and Children’s Hospital, Singapore; Cancer and Stem Cell Biology Program, Duke-NUS Graduate Medical School, Singapore
* These authors contributed equally to this work
Published in Cytotherapy 2016 18: 740-753 (Online Version)
There are efforts to differentiate human mesenchymal stem cells (hMSC) into cartilage tissue, for use in cell therapies. Conventional planar cultures are unable to provide the large cell numbers required to meet potential market demand for these therapeutic applications. Thus there is interest in using microcarrier-based cultures that are scalable for use in industrial-scale bioreactors. However, it remains unclear how microcarrier cultures affect hMSC chondrogenic potential. Here, we compared the chondrogenic potential of human early MSC (heMSC) between microcarrier-spinner and planar tissue culture plastic cultures.
heMSC were expanded in planar tissue-culture plastic cultures, or on collagen-coated Cytodex 3 microcarriers in spinner cultures. Chondrogenic pellets were generated from these cultures, and differentiated using various cytokine inducers of chondrogenesis. The extent of chondrogenic differentiation was investigated by measuring chondrogenic pellet diameter, DNA content, glycosaminoglycan (GAG) and Collagen II production, histological staining and gene expression of chondrogenic markers including Sox9, S100β, MMP13 and ALPL.
Our study revealed three main findings. First, BMP2 was the most effective chondrogenic inducer for heMSCs tested (compared to other cytokine inducers of chondrogenesis). Second, heMSC expanded in microcarrier-spinner cultures developed improved chondrogenic features, both functionally at the protein level and molecularly at the transcriptome level, compared to those of conventional static cultures on tissue culture plastic. Chondrogenic pellets generated from microcarrier cultures developed larger diameters, and produced more DNA, GAG and Collagen II per pellet with greater GAG/DNA and Collagen II/DNA ratios. Moreover, they induced higher expression of chondrogenic genes (e.g. S100β). Lastly, another microcarrier type showed similar trends in enhancing hMSC chondrogenic potential in microcarrier-spinner cultures suggesting that agitated nature of microcarrier cultures enhances chondregenesis.
In conclusion, we demonstrate that scalable microcarrier-spinner cultures enhance the chondrogenic potential of heMSC, supporting their use for large-scale cell expansion in cartilage cell therapy.